Search

Disturbances in acid-base balance, such as acidosis and alkalosis, have potential to alter the pharmacologic and toxicologic outcomes of statin therapy. Statins are commonly prescribed for elderly patients who have multiple comorbidities such as diabetes mellitus, cardiovascular, and renal diseases. These patients are at risk of developing acid-base imbalance. In the present study, the effect of disturbances in acid-base balance on the interconversion of simvastatin and pravastatin between lactone and hydroxy acid forms have been investigated in physiological buffers, human plasma, and cell culture medium over pH ranging from 6.8-7.8. The effects of such interconversion on cellular uptake and myotoxicity of statins were assessed in vitro using C2C12 skeletal muscle cells under conditions relevant to acidosis, alkalosis, and physiological pH. Results indicate that the conversion of the lactone forms of simvastatin and pravastatin to the corresponding hydroxy acid is strongly pH dependent. At physiological and alkaline pH, substantial proportions of simvastatin lactone (SVL; ∼87% and 99%, respectively) and pravastatin lactone (PVL; ∼98% and 99%, respectively) were converted to the active hydroxy acid forms after 24 hours of incubation at 37°C. At acidic pH, conversion occurs to a lower extent, resulting in greater proportion of statin remaining in the more lipophilic lactone form. However, pH alteration did not influence the conversion of the hydroxy acid forms of simvastatin and pravastatin to the corresponding lactones. Furthermore, acidosis has been shown to hinder the metabolism of the lactone form of statins by inhibiting hepatic microsomal enzyme activities. Lipophilic SVL was found to be more cytotoxic to undifferentiated and differentiated skeletal muscle cells compared with more hydrophilic simvastatin hydroxy acid, PVL, and pravastatin hydroxy acid. Enhanced cytotoxicity of statins was observed under acidic conditions and is attributed to increased cellular uptake of the more lipophilic lactone or unionized hydroxy acid form. Consequently, our results suggest that comorbidities associated with acid-base imbalance can play a substantial role in the development and potentiation of statin-induced myotoxicity., (Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.)

Interpretation of complex cancer genome data, generated by tumor target profiling platforms, is key for the success of personalized cancer therapy. How to draw therapeutic conclusions from tumor profiling results is not standardized and may vary among commercial and academically-affiliated recommendation tools. We performed targeted sequencing of 315 genes from 75 metastatic breast cancer biopsies using the FoundationOne assay. Results were run through 4 different web tools including the Drug-Gene Interaction Database (DGidb), My Cancer Genome (MCG), Personalized Cancer Therapy (PCT), and cBioPortal, for drug and clinical trial recommendations. These recommendations were compared amongst each other and to those provided by FoundationOne. The identification of a gene as targetable varied across the different recommendation sources. Only 33% of cases had 4 or more sources recommend the same drug for at least one of the usually several altered genes found in tumor biopsies. These results indicate further development and standardization of broadly applicable software tools that assist in our therapeutic interpretation of genomic data is needed. Existing algorithms for data acquisition, integration and interpretation will likely need to incorporate artificial intelligence tools to improve both content and real-time status., Competing Interests: No conflicts to disclose except for exceptions below: Eric Reiner: Consulting or Advisory Role: BSCI, Surefire Medical; Other Relationship: Sirtex Medical. Sarah Schellhorn Mougalian: Stock and Other Ownership Interests: Gilead Science, Coronado Biosciences, Roche; Research Funding: Genentech. Maysa M. Abu-Khalaf: Research Funding: Novartis, Merck, Clovis Oncology, Pfizer, Genentech/Roche, Merrimack. Lajos Pusztai: Consulting or Advisory Role: Clovis Oncology, Celgene; Honoraria: BioTheranostics, Pfizer; Research Funding: Foundation Medicine, Merck, Genentech.

Natural products play a pivotal role in medicine especially in the cancer arena. Many drugs that are currently used in cancer chemotherapy originated from or were inspired by nature. Jerantinine B (JB) is one of seven novel Aspidosperma indole alkaloids isolated from the leaf extract of Tabernaemontana corymbosa. Preliminary antiproliferative assays revealed that JB and JB acetate significantly inhibited growth and colony formation, accompanied by time- and dose-dependent apoptosis induction in human cancer cell lines. JB significantly arrested cells at the G2/M cell cycle phase, potently inhibiting tubulin polymerisation. Polo-like kinase 1 (PLK1; an early trigger for the G2/M transition) was also dose-dependently inhibited by JB (IC50 1.5 µM). Furthermore, JB provoked significant increases in reactive oxygen species (ROS). Annexin V+ cell populations, dose-dependent accumulation of cleaved-PARP and caspase 3/7 activation, and reduced Bcl-2 and Mcl-1 expression confirm apoptosis induction. Preclinical in silico biopharmaceutical assessment of JB calculated rapid absorption and bioavailability >70%. Doses of 8-16 mg/kg JB were predicted to maintain unbound plasma concentrations >GI50 values in mice during efficacy studies. These findings advocate continued development of JB as a potential chemotherapeutic agent., (Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.)

There has been increased interest in the medical use of cannabinoids in recent years, particularly in the predominant natural cannabinoids, cannabidiol (CBD) and Δ(9)-tetrahydrocannabinol (THC). The aim of the current study was to develop a sensitive and reliable method for the quantification of CBD and THC in rat plasma. A combination of protein precipitation using cold acetonitrile and liquid-liquid extraction using n-hexane was utilised to extract CBD and THC from rat plasma. Samples were then evaporated and reconstituted in acetonitrile and 30 μL was injected into an HPLC system. Separation was achieved using an ACE C18-PFP 150 mm × 4.6 mm, 3 μm column at 55 °C with isocratic elution using a mobile phase consisting of acetonitrile-water (62:38, v/v) at 1 mL/min for 20 min. Both cannabinoids, as well as the internal standard (4,4-dichlorodiphenyltrichloroethane, DDT) were detected at 220 nm. Our new method showed linearity in the range of 10-10,000 ng/mL and a lower limit of quantification (LLOQ) of 10 ng/mL for both cannabinoids, which is comparable to previously reported LC-MS/MS methods. Inter- and intra-day precision and accuracy were below 15% RSD and RE, respectively. To demonstrate the suitability of the method for in vivo studies in rats, the assay was applied to a preliminary pharmacokinetic study following IV bolus administration of 5 mg/kg CBD or THC. In conclusion, a simple, sensitive, and cost-efficient HPLC-UV method for the simultaneous determination of CBD and THC has been successfully developed, validated and applied to a pharmacokinetic study in rats., (Copyright © 2015 Elsevier B.V. All rights reserved.)

Purpose: Benzodiazepines (BDZs) are the drugs of choice to prevent the symptoms of alcohol withdrawal syndrome (AWS). Various treatment protocols are published and have been shown to be effective in both office-managed and facility-managed treatment of AWS. The aim of this scientific commentary is to demonstrate the differences in the expected exposure to BDZs during AWS treatment using different treatment regimens available in the literature, in patients with or without alcoholic liver cirrhosis., Methods: Diazepam and lorazepam AWS protocols were examined and reviewed in the literature, and blood plasma levels were examined and compared, respectively., Results: Considerable variation in the blood levels with the different dosing schedules was found. Because the drugs are metabolized differently, we have also shown that liver disease affects the blood levels of diazepam, but not of lorazepam., Conclusions: Differences in treatment regimens, the choice of BDZ, as well as the presence of liver cirrhosis can substantially alter the exposure of patients to drugs used for AWS treatment. Outpatient treatment of AWS has been shown to be relatively safe and effective for the treatment of AWS but patients should be carefully monitored.

Triglycerides (TG) are one of the most common excipients used in oral lipid-based formulations. The chain length of the TG plays an important role in the oral bioavailability of the co-administered drug. Fatty acid (FA) chain-length specificity of porcine pancreatic lipase was studied by means of an in vitro lipolysis model under bio-relevant conditions at pH 6.80. In order to determine the total extent of lipolysis, back-titration experiments at pH 11.50 were performed. Results suggest that there is a specific chain length range (C2-C8) for which pancreatic lipase shows higher activity. This specificity could result from a combination of physicochemical properties of TGs, 2-monoglycerides (2-MGs) and FAs, namely the droplet size of the TGs, the solubility of 2-MGs within mixed micelles, and the relative stability of the FAs as leaving groups in the hydrolysis reaction. During experimentation, it was evident that an optimisation of lipolysis conditions was needed for tighter control over pH levels so as to better mimic in vivo conditions. 1M NaOH, 3.5 mL/min maximum dosing rate, and 3 μL/min minimum dosing rate were the optimised set of conditions that allowed better pH control, as well as the differentiation of the lipolysis of different lipid loads., (Copyright © 2015 Elsevier B.V. All rights reserved.)

Human saliva is a biological fluid of great importance in the field of dissolution testing. However, until now, no consensus has been reached on its key characteristics relevant to dissolution testing. As a result, it is difficult to select or develop an in vitro dissolution medium to best represent human saliva. In this study, the pH, buffer capacity, surface tension, viscosity and flow rate of both unstimulated (US) and stimulated (SS) human saliva were investigated in order to provide a platform of reference for future dissolution studies using simulated salivary fluids. Age and gender related differences in a sample size of 30 participants for each parameter were investigated. Significant differences were established between US and SS for all characteristics except surface tension. Therefore, the requirement for using two simulated salivary fluids should be considered when developing an oral dissolution model., (Copyright © 2015 Elsevier B.V. All rights reserved.)

Consumption of plant sterols/ stanols has long been demonstrated to reduce plasma cholesterol levels. The objective of this review is to demonstrate the lipid-lowering activity and anti-atherogenic effects of natural and semi-synthetic plant sterols/ stanols based on evidence from cell-culture studies, animal studies and clinical trials. Additionally, this review highlights certain molecular mechanisms by which plant sterols/ stanols lower plasma cholesterol levels with a special emphasis on factors that affect the cholesterol-lowering activity of plant sterols/stanols. The crystalline nature and the poor oil solubility of these natural products could be important factors that limit their cholesterol-lowering efficiency. Several attempts have been made to improve the cholesterol-lowering activity by enhancing the bioavailability of crystalline sterols and stanols. Approaches involved reduction of the crystal size and/or esterification with fatty acids from vegetable or fish oils. However, the most promising approach in this context is the chemical modification of plant sterols /stanols into water soluble disodium ascorbyl phytostanyl phosphates analogue by esterification with ascorbic acid. This novel semi-synthetic stanol derivative has improved efficacy over natural plant sterols/ stanols and can provide additional benefits by combining the cholesterol-lowering properties of plant stanols with the antioxidant potential of ascorbic acid. This article is open to POST-PUBLICATION REVIEW. Registered readers (see "For Readers") may comment by clicking on ABSTRACT on the issue's contents page.

Purpose: To evaluate the antifungal activity of amphotericin B (AmB) in a mouse model of systemic candidiasis following administration of a novel oral AmB formulation (iCo-010) that has been pre-exposed to tropical temperatures., Methods: Amphotericin B (AmB) was prepared as a 5 mg/mL dispersion in a mixture of Peceol, Gelucire 44/14 and VitE-TPGS 2,3 (iCo-010). The formulation was protected from light and incubated in a sealed container at 43 °C for 60 days. Mice infected with Candida albicans were treated with either iCo-010 formulation pre-incubated at 43 °C for 60 days or freshly prepared iCo-010 formulation at doses of 5, 10 and 20 mg/kg once daily for five consecutive days. Single intravenous 5 mg/kg dose of AmBisome® was used as a positive control group. Seven days following the last dose, the kidney, liver, spleen, lung, heart and brain were removed and the number of colony forming units (CFUs) was determined as a measure of tissue fungal load. In addition, the concentration of AmB within each tissue was determined using high performance liquid chromatography (HPLC)., Results: There were no significant differences in the reduction of CFUs and the concentration of AmB recovered in all organs at all iCo-010 doses tested between the freshly prepared iCo-010 formulation compared to the formulation that was incubated at 43 °C for 60 days., Conclusions: A novel oral AmB formulation, iCo-010, incubated at 43 °C for 60 days to simulate the exposure of the formulation to tropical temperatures remained highly effective against murine systemic candidiasis.

Neglected tropical diseases (NTDs) are infections which are endemic in poor populations in lower- and middle-income countries (LMIC). Approximately one billion people have now or are at risk of getting an NTD and yet less than 5% of research dollars are focused on providing treatments and prevention of these highly debilitating and deadly conditions. The United States Food and Drug Administration (FDA) Orphan Drug Designation program (ODDP) provides orphan status to drugs and biologics, defined as those intended for the safe and effective treatment, diagnosis or prevention of rare diseases and/or disorders that affect fewer than 200 000 people in the United States, or that affect more than 200 000 persons but are not expected to recover the costs of developing and marketing a treatment drug. These regulations have led to the translation of rare disease knowledge into innovative rare disease therapies. The FDA Guidance for Industry on developing drugs for the treatment and prevention of NTDs describes the following regulatory strategies: Orphan Product Designation, Fast Track Designation, Priority Review Designation, Accelerated Approval and Tropical Disease Priority Review Voucher. This paper will discuss how these regulations and especially the ODDP can improve the clinical development and accessibility of drug products for NTDs.

Statins are lipid-lowering drugs used widely to prevent and treat cardiovascular and coronary heart diseases. These drugs are among the most commonly prescribed medicines intended for long-term use. In general, statins are well tolerated. However, muscular adverse effects appear to be the most common obstacle that limits their use, resulting in poor patient compliance or even drug discontinuation. In addition, rare but potentially fatal cases of rhabdomyolysis have been reported with the use of these drugs, especially in the presence of certain risk factors. Previous reports have investigated statin-induced myotoxicity in vivo and in vitro using a number of cell lines, muscle tissues, and laboratory animals, in addition to randomized clinical trials, observational studies, and case reports. None of them have compared directly results from laboratory investigations with clinical observations of statin-related muscular adverse effects. To the best of our knowledge this is the first review article that combines laboratory investigation with clinical aspects of statin-induced myotoxicity. By reviewing published literature of in vivo, in vitro, and clinically relevant studies of statin myotoxicity, we aim to translate this important drug-related problem to establish a clear picture of proposed mechanisms that explain the risk factors and describe the diagnostic approaches currently used for evaluating the degree of muscle damage induced by these agents. This review provides baseline novel translational insight that can be used to enhance the safety profile, to minimize the chance of progression of these adverse effects to more severe and potentially fatal rhabdomyolysis, and to improve the overall patient compliance and adherence to long-term statin therapy., (Copyright © 2014 Mosby, Inc. All rights reserved.)

Objectives: Amended reports (AmRs) need to follow patients to treating physicians, to avoid erroneous management based on the original diagnosis. This study was undertaken to determine if AmRs followed the patient appropriately., Methods: AmRs with diagnostic changes and discrepancies between ordering and treating physicians were tracked. Chart reviews, electronic medical report (EMR) reviews, and interviews were conducted to establish receipt of the AmR by the correct physician., Results: Seven of 60 AmRs had discrepancies between the ordering and treating physicians, all with malignant diagnoses. The AmR was present in the treating physician's chart in only one case. Ordering physicians indicated that AmRs were not forwarded to treating physicians when corrected results arrived after patient referral, under the assumption that the new physician was automatically forwarded pathology updates. No harm was documented in any of our cases. In one case with a significant amendment, the correct information was entered in the patient chart based on a tumor board discussion. A review of two electronic health record systems uncovered significant shortcomings in each delivery system., Conclusions: AmRs fail to follow the patient's chain of referrals to the correct care provider, and EMR systems lack the functionality to address this failure and alert clinical teams of amendments., (Copyright© by the American Society for Clinical Pathology.)

Purpose: To develop and to validate a simple but sensitive method for determination of vitamins D3 and K1 in rat plasma., Methods: The sample treatment included protein precipitation by cold acetonitrile, evaporation, reconstitution with methanol and filtration. The chromatography conditions included Xterra RP18 3.5 µm 4.6 × 100 mm column at ambient temperature and mobile phase consisting of methanol/water (93/7, v/v) at 0.5 mL/min flow rate. Vitamin D3 and probucol were detected at 265 nm and vitamin K1 at 239 nm. Rats were administered intravenously by 0.1 mg/kg of vitamin D3 or K1 and the blood samples were withdrawn pre-administration and at pre-determined time points post-administration. The pharmacokinetic analysis was performed using a non-compartmental approach., Results: The calibration curves in rat plasma were linear up to 5000 ng/mL for both vitamins. The limit of quantification (LOQ) was 20 ng/mL for vitamin D3 and 40 ng/mL for K1. Inter- and intra-day precision and accuracy were below 15%. The pharmacokinetic parameters of vitamin D3 following intravenous administration were: AUC0-∞ = 11323 ± 1081 h × ng/mL, Vd = 218 ± 80 mL/kg, CL = 8.9 ± 0.8 mL/h/kg, t1/2 = 16.8 ± 5 h; and of vitamin K1: AUC0-∞ = 2495 ± 297 h × ng/mL, Vd = 60 ±24 mL/kg, CL = 40.5 ± 5.1 mL/h/kg, t1/2 = 1.1 ±0.5 h., Conclusion: The developed HPLC-UV assay is a simple and sensitive method for the determination of vitamins D3 and K1 in rat plasma. A higher dose of vitamin K1 should be used in future studies for accurate estimation of pharmacokinetic parameters. The data show the suitability of the assay for pharmacokinetic studies in rats.

Context: In 2007 the American Society of Clinical Oncology/College of American Pathologists made new recommendations for HER2 testing and redefined HER2 positivity., Objective: To analyze results from simultaneous HER2 testing with immunohistochemistry and fluorescence in situ hybridization (FISH) in 2590 invasive breast carcinomas between 2002 and 2010, using 2 scoring systems., Design: Cases from between 2002 and 2006 were scored by using original US Food and Drug Administration criteria (N = 1138) and those from between 2007 and 2010 were evaluated according to American Society of Clinical Oncology/College of American Pathologists criteria (N = 1452). Concordance between testing methods and clinicopathologic associations were determined., Results: Overall concordance between immunohistochemistry/FISH in the 9-year period was 96.2% (κ = 0.82), and positive concordance was lower. After 2007, the proportion of HER2/neu-positive and HER2/neu-negative cases was not significantly changed when using immunohistochemistry (10.5% versus 8.9%, P = .22 and 69.4% versus 63%, P = .13, respectively), but the number of equivocal cases was higher (19.9% versus 28%, P < .001). While the proportion of negative cases by FISH remained unchanged after 2007 (86.5% versus 88.2%, P = .76), the number of positive cases was lower (13.4% versus 9.2%, P < .001). In addition, 38 cases (2.6%) were FISH equivocal, 16 of which were also equivocal by immunohistochemistry. Overall, immunohistochemistry/FISH concordance was 95.9% between 2002 and 2006 (κ = 0.82) and 96.4% after 2007 (κ = 0.82). However, an approximately 13% lower positive assay concordance was noted in the last period., Conclusions: Application of American Society of Clinical Oncology/College of American Pathologists recommendations is associated with comparable overall immunohistochemistry/FISH concordance, reduced positive concordance, and increased equivocal results.

Conventional adult dosage forms are often not suitable for the paediatric and geriatric populations due to either swallowing difficulties or patient repulsion and a requirement for tailored dosing to individual compliance or physiological needs. Alternative formulations are available; however these often require the incorporation of more complex taste masking techniques. One approach to taste masking is to reduce contact between the bitter Active Pharmaceutical Ingredient (API) and oral cavity taste bud regions. This is achieved by hindering release in the oral cavity, or including competitive inhibition of bitter sensation for example by using flavours or sweeteners. There may also be other sensational complications from the API such as residual burning, reflux or metallic taste sensations to deal with. In vitro dissolution testing is employed to elucidate taste masking capability by quantifying release of the drug in simulated oral cavity conditions. Dissolution testing approaches may also be used to potentially predict or quantify the effect of the taste masking technique on the resultant pharmacokinetic profile. The present review investigates the anatomy and physiology of the oral cavity and current approaches to taste masking. In vitro dissolution methodologies adopted in the evaluation of taste masked formulations are discussed for their relative merits and drawbacks. A vast array of methodologies has been employed, with little agreement between approaches, and a lack of biorelevance. Future directions in dissolution methodology such as TNO Intestinal Model (TIM) and the Artificial Stomach and Duodenum model (ASD) are also discussed., (© 2013.)

Purpose: To investigate the biodistribution of amphotericin B (AmB) in mice and rats following administration of liposomal AmB (AmBisome®) using a physiologically-based pharmacokinetic (PBPK) modeling framework and to utilize this approach for predicting AmBisome® pharmacokinetics in human tissues., Methods: AmB plasma and tissue concentration-time data, following single and multiple intravenous administration of nonliposomal and liposomal AmB to mice and rats, were extracted from literature. The whole-body PBPK model was constructed and incorporated nonliposomal and liposomal subcompartments. Various structural models for individual organs were evaluated. Allometric relationships were incorporated into the model to scale parameters based on species body weight., Results: A non-Michaelis-Menten mechanism was included into the structure of the liver and spleen liposomal compartments to describe saturable uptake of particles by the reticuloendothelial system. The model successfully described plasma and tissue pharmacokinetics of AmB after administration of AmBisome® to rats and mice., Conclusions: The dual PBPK model demonstrated good predictive performance by reasonably simulating AmB exposure in human tissues. This modeling framework can be potentially utilized for optimizing AmBisome® therapy in humans and for investigating pathophysiological factors controlling AmB pharmacokinetics and pharmacodynamics.

The objective of this study was to assess the pharmacokinetics and tissue distribution of amphotericin B (AmB) in rats following oral administration of three lipid-based formulations (iCo-009, iCo-010 and iCo-011). The lipid-based formulations were administered to rats at a dose of 10 mg/kg and blood samples were withdrawn at predose, 1, 2, 4, 6, 8, 10, 12, 24, 48 and 72 h, after which the animals were sacrificed and the body organs were collected for AmB quantification using a validated HPLC method. Plasma pharmacokinetics parameters were determined using non-compartmental analysis. The disappearance of AmB from plasma was the slowest following the administration of iCo-010 with MRT of 63 h followed by iCo-009 then iCo-011 (36 and 27 h). The AUC(0-24h) of iCo-009 and iCo-010 was 1.5-2-fold higher than that of iCo-011. The kidney exposure was comparable between iCo-009 and iCo-010 and was higher than that of iCo-011. The lung exposure was the highest following iCo-010 administration as compared to that of iCo-009. The distribution of AmB from plasma to tissues resulted in a high accumulation of AmB overtime with slow back-distribution to plasma. The pharmacokinetics profiles varied among the three formulations, despite the similarity in lipid composition between iCo-010 and iCo-011 and the presence of Peceol® as a common component in the formulations. The administration of oral iCo-010 could lead to higher steady state concentrations in the tissues after multiple dosing, which could lead to enhanced eradication of tissue borne fungal and parasitic infections.

Atherosclerosis, the gradual formation of a lipid-rich plaque in the arterial wall is the primary cause of Coronary Artery Disease (CAD), the leading cause of mortality worldwide. Hypercholesterolemia, elevated circulating cholesterol, was identified as a key risk factor for CAD in epidemiological studies. Since the approval of Mevacor in 1987, the primary therapeutic intervention for hypercholesterolemia has been statins, drugs that inhibit the biosynthesis of cholesterol. With improved understanding of the risks associated with elevated cholesterol levels, health agencies are recommending reductions in cholesterol that are not achievable in every patient with statins alone, underlying the need for improved combination therapies. The whole body cholesterol pool is derived from two sources, biosynthesis and diet. Although statins are effective at reducing the biosynthesis of cholesterol, they do not inhibit the absorption of cholesterol, making this an attractive target for adjunct therapies. This report summarizes the efforts to target the gastrointestinal absorption of cholesterol, with emphasis on specifically targeting the gastrointestinal tract to avoid the off-target effects sometimes associated with systemic exposure.

An in vitro lipolysis model was utilized to study the effect of stigmastanol (lipophilic phytosterol) and disodium ascorbyl phytostanol phosphate (DAPP) (modified hydrophilic phytostanol) on intestinal processing of cholesterol to gain further understanding of their cholesterol lowering mechanism. Lipolysis results showed that stigmastanol, if given in powder alone, had no effect on cholesterol processing probably due to its poor solubility. Stigmastanol suspension formulation re-distributed cholesterol from aqueous phase to oil and sediment phases. The water soluble DAPP has changed cholesterol distribution even more significantly by transferring cholesterol from aqueous phase to sediment phase. Moreover, the results provided evidence that DAPP inhibited triglyceride digestion in vitro. Considering DAPP as a surfactant with the same lipophilic sterol ring as bile salt, its ability to inhibit triglyceride lipolysis may be due to its competition with bile salt for the substrate surface, thereby hindering the lipolysis of triglyceride and inhibiting cholesterol solubilization with the lipolysis products. It can be speculated that the cholesterol lowering mechanism of DAPP during intestinal digestion is related to its ability to act as a surfactant closely resembling bile salt., (Copyright © 2012 Elsevier B.V. All rights reserved.)

The objective of this work was to assess the antifungal activity of a tropically stable formulation of amphotericin B (AmB) (iCo-010) over short period of treatment in a rat model of invasive candidiasis. The rats were infected with Candida albicans (ATCC 18804); 48 h later, the animals were assigned either to a control group, AmBisome(®) group (5 mg/kg QD), or iCo-010 groups (0.5, 1, 2.5, 5 and 10 mg/kg TID). The animals were treated for two days and then sacrificed 18 h following the completion of the treatment. The blood, liver, lungs, kidneys and spleen were harvested to assess the colony forming units in the samples. There was no significant difference in the reduction of the fungal burden in the organs between the AmBisome(®) and iCo-010 groups except in the spleen and liver. There was a linear correlation between the antifungal activity in renal tissues and the administered doses of iCo-010. The plasma creatinine levels were not significantly different among the control and all the treatment groups. Oral iCo-010 has high efficacy against invasive candidiasis in renal and pulmonary tissues. Longer treatment period than the two-days regimen should be considered for higher therapeutic efficacy of iCo-010 in all the tissues., (Copyright © 2012 Elsevier B.V. All rights reserved.)

The purpose of this study was to investigate the intraluminal processing of novel oral lipid-based formulations of amphotericin B using an in vitro lipolysis model. Amphotericin B (AmB) was formulated in three lipid-based formulations consisting of different lipid components: iCo-009, iCo-010 and iCo-011. Various lipid loads (0.25, 0.5, 1 and 2 g) were digested using the lipolysis model to assess AmB distribution among the lipolysis phases. The duration of lipolysis was comparable among the three formulations except for 2 g load of iCo-009 which had a significantly longer lipolysis than iCo-010 and iCo-011. The lipid components of iCo-009 experienced lower extent of lipolysis as compared to other formulations. Amphotericin B concentration in the aqueous phases was the highest with iCo-010 which also had the lowest sediment recovery. Amphotericin B levels in the undigested lipid layers were comparable between iCo-009 and iCo-010 and were higher than with iCo-011. Given the observation that iCo-010 had the highest aqueous micellar solubilization and the lowest sediment recovery of AmB among the tested formulations, these results could potentially be used to interpret and predict the in vivo performance of AmB- SEDDS formulations in future studies., (Crown Copyright © 2012. Published by Elsevier B.V. All rights reserved.)

Purpose: Currently, in vivo or in vitro(99m)Tc-radiolabelled red blood cells are the standard blood pool imaging agents. Due to risks associated with handling of blood and the problems with the current (99m)Tc shortage, we were interested in a long-circulating biocompatible synthetic macromolecule that would be simple to prepare and could also be used for PET imaging., Methods: A high molecular weight hyperbranched polyglycerol (HPG) of 500 kDa was derivatized to coordinate radioactive gallium and to establish its labelling efficiency, stability and pharmacokinetics., Results: The resulting radiopharmaceutical in kit form was labelled rapidly within a couple of minutes at room temperature, was stable in transferrin and EDTA challenge tests, and was non-toxic in both cell viability and different hemocompatibility assays. A pharmacokinetic biodistribution study showed that the (67)Ga-HPGN was confined to the blood compartment with a biological half life of 50.7h., Conclusion: (67)Ga-HPGN is thus a simple to prepare blood pool imaging agent for applications where a long biological half-life is essential, i.e., the diagnosis of internal bleeding. Since radiolabelling of the same kit with (68)Ga was also confirmed, we plan to evaluate it shortly as a PET blood pool imaging agent for cardiac applications., (Copyright © 2011 Elsevier B.V. All rights reserved.)

Protonated nanostructured aluminosilicate (NSAS) was previously reported as effective in decreasing intestinal absorption of cholesterol in rats. In this work, the effect of NSAS on modeled intraluminal distribution of cholesterol was assessed using an in vitro lipolysis model. In addition to NSAS, the effect of bile salt sequestrant cholestyramine on modeled intraluminal distribution of cholesterol was tested, as well as the effect of the absence of bile salts in the system. NSAS induced sharp redistribution of cholesterol from aqueous to sediment phase, which suggests that NSAS exerts its inhibition of cholesterol absorption by direct or indirect binding to cholesterol, followed by its precipitation and excretion with feces. Cholestyramine induced redistribution of cholesterol into both oil and sediment phase. The redistribution into the sediment phase in the presence of cholestyramine was more intensive than when bile acids were omitted from the system, which suggests potential cholesterol-lowering activity of this compound on the level of intraluminal processing of cholesterol. Assessment of intraluminal processing of cholesterol using in vitro dynamic lipolysis model can potentially become a valuable approach to assess the action of cholesterol absorption inhibitors in a fast and efficient manner., (Copyright © 2011 Wiley-Liss, Inc.)

In-house software development for use in a clinical laboratory is a controversial issue. Many of the objections raised are based on outdated software development practices, an exaggeration of the risks involved, and an underestimation of the benefits that can be realized. Buy versus build analyses typically do not consider total costs of ownership, and unfortunately decisions are often made by people who are not directly affected by the workflow obstacles or benefits that result from those decisions. We have been developing custom software for clinical use for over a decade, and this article presents our perspective on this practice. A complete analysis of the decision to develop or purchase must ultimately examine how the end result will mesh with the departmental workflow, and custom-developed solutions typically can have the greater positive impact on efficiency and productivity, substantially altering the decision balance sheet. Involving the end-users in preparation of the functional specifications is crucial to the success of the process. A large development team is not needed, and even a single programmer can develop significant solutions. Many of the risks associated with custom development can be mitigated by a well-structured development process, use of open-source tools, and embracing an agile development philosophy. In-house solutions have the significant advantage of being adaptable to changing departmental needs, contributing to efficient and higher quality patient care.

Background: Invasive fungal infections such as candidiasis constitute an increasingly important medical problem. Drugs currently used for the treatment of candidiasis include polyenes (such as Amphotericin B) and azoles. Amphotericin B (AmpB) presents several limitations such as its nephrotoxicity and limited solubility. We have developed two novel lipid-based AmpB formulations which in vivo show less nephrotoxicity and enhanced solubility compared to Fungizone™ a commercial AmpB formulation. The purpose of this study was to determine the cytotoxicity of Fungizone™, Ambisome™ and two novel AmpB formulations (iCo-009 and iCo-010) against Candida albicans, human kidney (293T) cells and monocytic (THP1) cells., Methods: Cell cytotoxicity to the AmpB formulations was evaluated by MTS and LDH assays. In vitro anti-Candida albicans activity was assessed after a 48 h drug incubation., Results: None of the AmpB formulations tested showed cytotoxicity against 293T cells. In the case of THP1 cells only Fungizone™ and Ambisome™ showed cytotoxicity at 500 μg/L (n = 4-10, p < 0.05).The calculated EC50 to Candida albicans for the different formulations was as follows: 26.8 ± 2.9 for iCo-010, 74.6 ± 8.9 for iCo-009, 109 ± 31 for Ambisome™ and 87.1 ± 22 for Fungizone™ (μg of AmpB/L, n = 6-12, p < 0.05)., Conclusions: The AmpB formulations analyzed were not cytotoxic to 293T cells. Cytotoxicity in THP1 cells was observed for Fungizone™ and Ambisome™, but not with the novel AmpB formulations. iCo-010 had higher efficacy compared to other three AmpB formulations in the Candida albicans model.The absence of cytotoxicity as well as its higher efficacy for the Candida model compared to Fungizone™ and Ambisome™ suggest that iCo-010 has potential in treating candidiasis.

Background: The purpose of this study was to evaluate the biodistribution and toxicity of amphotericin B (AmB) following multiple oral administrations of a novel tropically stable lipid-based formulation (iCo-010)., Methods: BALB/c mice were allocated into six groups: oral iCo-010 twice daily for 5 days in the dose of 20, 10, 5 and 2.5 mg/kg; vehicle control; and intravenous boluses of Fungizone 2 mg/kg once daily for 5 days. The animals were sacrificed 12 h following the last administration and blood and tissues were collected., Results: The plasma concentrations of AmB were similar to previously reported after administration of iCo-009. Somewhat lower concentrations of AmB were detected in reticulo-endothelial system in the case of iCo-010 when compared with iCo-009. The concentration in kidney was higher with iCo-010 than with iCo-009. The creatinine levels in all oral treatment groups were in a normal range as in the case of iCo-009. Administration of Fungizone resulted in elevated plasma creatinine levels. Histopathology analysis detected no GI, liver or kidney toxicity following multiple dose oral administration of iCo-010. Fungizone treatment induced necrotic changes in hepatic and kidney tissues., Conclusions: Given the tropical stability of iCo-010, near identical activity against visceral leishmaniasis and significant concentrations in target organs this formulation has a potential to become a treatment of choice in tropical developing countries.

Purpose: Recent misclassification (false negative) incidents have raised awareness concerning limitations of immunohistochemistry (IHC) in assessment of estrogen receptor (ER) in breast cancer. Here we define a new method for standardization of ER measurement and then examine both change in percentage and threshold of intensity (immunoreactivity) to assess sources for test discordance., Methods: An assay was developed to quantify ER by using a control tissue microarray (TMA) and a series of cell lines in which ER immunoreactivity was analyzed by quantitative immunoblotting in parallel with the automated quantitative analysis (AQUA) method of quantitative immunofluorescence (QIF). The assay was used to assess the ER protein expression threshold in two independent retrospective cohorts from Yale and was compared with traditional methods., Results: Two methods of analysis showed that change in percentage of positive cells from 10% to 1% did not significantly affect the overall number of ER-positive patients. The standardized assay for ER on two Yale TMA cohorts showed that 67.9% and 82.5% of the patients were above the 2-pg/μg immunoreactivity threshold. We found 9.1% and 19.7% of the patients to be QIF-positive/IHC-negative, and 4.0% and 0.4% to be QIF-negative/IHC-positive for a total of 13.1% and 20.1% discrepant cases when compared with pathologists' judgment of threshold. Assessment of survival for both cohorts showed that patients who were QIF-positive/pathologist-negative had outcomes similar to those of patients who had positive results for both assays., Conclusion: Assessment of intensity threshold by using a quantitative, standardized assay on two independent cohorts suggests discordance in the 10% to 20% range with current IHC methods, in which patients with discrepant results have prognostic outcomes similar to ER-positive patients with concordant results.

Simulation of microgravity for cultivated multipotent mesenchymal stromal cells (MSCs) from human marrow changes transiently expression of genes associated with actin cytoskeleton; the effect fades away partially in 120 hrs. following microgravity and completely after 24-hr cell readaptation to static conditions. These changes in expression of some cytoskeleton genes seem to predetermine their reaction to simulated microgravity, and therefore inhibition of MSCs differentiation potential.

The time course of tissue distribution of amphotericin B (AmB) has not been sufficiently characterized despite its therapeutic importance and an apparent disconnect between plasma pharmacokinetics and clinical outcomes. The goals of this work were to develop and evaluate a physiologically based pharmacokinetic (PBPK) model to characterize the disposition properties of AmB administered as deoxycholate formulation in healthy rats and to examine the utility of the PBPK model for interspecies scaling of AmB pharmacokinetics. AmB plasma and tissue concentration-time data, following single and multiple intravenous administration of Fungizone® to rats, from several publications were combined for construction of the model. Physiological parameters were fixed to literature values. Various structural models for single organs were evaluated, and the whole-body PBPK model included liver, spleen, kidney, lung, heart, gastrointestinal tract, plasma, and remainder compartments. The final model resulted in a good simultaneous description of both single and multiple dose data sets. Incorporation of three subcompartments for spleen and kidney tissues was required for capturing a prolonged half-life in these organs. The predictive performance of the final PBPK model was assessed by evaluating its utility in predicting pharmacokinetics of AmB in mice and humans. Clearance and permeability-surface area terms were scaled with body weight. The model demonstrated good predictions of plasma AmB concentration-time profiles for both species. This modeling framework represents an important basis that may be further utilized for characterization of formulation- and disease-related factors in AmB pharmacokinetics and pharmacodynamics.

Purpose: To develop an HPLC-UV method for determination of a novel antitrypanosomal compound (OSU-36) and its ester prodrug (OSU-40) in rat plasma and to apply the method for pharmacokinetic evaluation of both compounds in rats., Methods: Since an attempt to assay for OSU-36 and OSU-40 in non-stabilized plasma resulted in highly non-linear calibration curves and poor sensitivity due to instability of the compounds, the plasma was stabilized using paraoxon and ascorbic acid. The sample treatment included protein precipitation by acetonitrile; evaporation; reconstitution with acetonitrile and filtration. The chromatography conditions included Xterra RP18 3.5 µm 4.6X100 mm column and gradient mobile phase system of acetonitrile-water., Results: The limits of quantification (LOQ) were 50 ng/mL and 40 ng/mL for OSU-36 and OSU-40, respectively. The intra- and interday precision and accuracies were below 13% for low, medium and high quality control samples for both compounds. While OSU-40 has been stable in all tested handling conditions, OSU-36 was unstable in plasma after 20 days storage in -80 °C or 4h 28 °C storage. The developed method has been applied for a pharmacokinetic study in rats which revealed that an ester prodrug OSU-40 is rapidly converted to OSU-36 within the plasma compartment by plasma esterases. OSU-36, in turn, relatively quickly undergoes oxidative metabolism, including within the plasma compartment., Conclusions: A supplementation of rat plasma with an esterase inhibitor to prevent degradation of ester prodrug (OSU-40), and with antioxidant to prevent oxidation of OSU-36, is necessary for reliable determination of both compounds. Due to limited stability of OSU-36 in stabilized rat plasma, long-term storage of samples or prolonged handling in room temperature conditions is not recommended.

Analogs of the trypanocidal lead compound 1-benzyl-1,2-dihydro-2,2,4-trimethylquinolin-6-yl acetate were prepared to extend the structure-activity relationship in this series of molecules, improve the in vivo antitrypanosomal activity of the lead, and determine whether ester prodrugs are needed to overcome the instability of the dihydroquinolin-6-ols. Two of the most active compounds identified in this study were 1-benzyl-1,2-dihydro-2,2,4-trimethylquinolin-6-ol hydrochloride and 1-(2-methoxy)benzyl-1,2-dihydro-2,2,4-trimethylquinolin-6-ol hydrochloride. These stable solids possessed low nanomolar IC(50) values against Trypanosoma brucei rhodesiense STIB900 in vitro and provided cures in an early treatment acute mouse model of African trypanosomiasis when given ip at 50mg/kg/day for four consecutive days., (Copyright © 2010 Elsevier Ltd. All rights reserved.)

Purpose: To develop an oral formulation of amphotericin B (AmB) that is stable at the temperatures of WHO Climatic Zones 3 and 4 (30-43 °C) and to evaluate its efficacy in a murine model of visceral leishmaniasis (VL)., Methods: The stability testing of four novel oral lipid AmB formulations composed of mono- and di-glycerides and pegylated esters (iCo-010 to iCo-013) was performed over 60 d and analyzed by HPLC-UV. In addition, the four formulations were incubated 4 h in fasted-state simulated intestinal fluid. AmB concentration was measured spectrophotometrically and emulsion droplet diameter was assessed by dynamic light scattering. Antileishmanial activity of iCo-010 was evaluated at increasing oral doses (2.5 to 10 mg/kg) in a murine model of VL., Results: AmB stability in the lipid formulation (iCo-010) was >75% over 60 days. After 4 h in fasted-state simulated intestinal fluid, AmB concentration was >95%. iCo-010 demonstrated significant efficacy when orally administered to VL-infected mice bid for five days (inhibition of 99%, 98%, and 83% at 10, 5 and 2.5 mg/kg compared to the vehicle control). In addition, the qd dose of 20 mg/kg provided 96% inhibition compared to the vehicle control., Conclusions: The oral AmB formulation iCo-010 is stable at the temperatures of WHO Climatic Zones 3 and 4 (30-43 °C). iCo-010 showed excellent antileishmanial activity at both 10 mg/kg po bid for 5 days (<99% reduction in parasitic infection) and 20 mg/kg po qd for 5 days (95% inhibition when compared to control).

There is a long-term discussion in the literature concerning the possible link between the improved efficacy of clozapine treatment and elevated plasma triglyceride levels, but no mechanistic studies have been performed to date. The aim of this work was to investigate whether the postprandial hypertriglyceridemia affects the pharmacokinetics and brain distribution of clozapine and norclozapine. Experimental hypertriglyceridemia in rats was induced by oral administration of peanut oil and the pharmacokinetic parameters and brain penetration of clozapine and norclozapine following administration of clozapine were compared to normotriglyceridemic control animals. Moderately increased clearance of clozapine was found in hypertriglyceridemic animals compared to control group. No changes were found in penetration of compounds across the blood-brain barrier (BBB). Taken together, the results do not support the hypothesis that hypertriglyceridemia improves the effect of clozapine by altered pharmacokinetics of clozapine and norclozapine and their increased penetration across the BBB., (Copyright 2010 Elsevier B.V. All rights reserved.)

Objectives: To assess the impact of visceral leishmaniasis (VL) on the concentration of amphotericin B (AmB) recovered in the liver and spleen following either intravenous (AmBisome) or oral (iCo-009) AmB administration to mice., Methods: Livers and spleens previously obtained from VL-infected BALB/c mice (following intravenous AmBisome or oral AmB treatments) were analysed for AmB concentrations. Then, non-infected BALB/c mice were divided into three treatment groups: a single dose of intravenous AmBisome (2 mg/kg, n = 5); and oral AmB every 12 h for 5 days (10 mg/kg, n = 6 and 20 mg/kg, n = 6). The animals were sacrificed 7 days after the initiation of the treatment and the livers and spleens were harvested for drug analysis by HPLC., Results: The single intravenous injection of AmBisome resulted in a 77-fold lower concentration of AmB in infected compared with non-infected liver tissue, while the difference in AmB concentration in the spleen was only 5-fold. The multiple dose oral administration of AmB resulted in a 3-fold lower concentration of AmB in infected compared with non-infected livers for both oral doses, while the differences in AmB concentrations in the spleen were not statistically different for the oral treatment groups., Conclusions: VL significantly lowered the concentration of AmB in the liver and the spleen when compared with uninfected animals. This effect seems to correlate with the degree of infection of the tissue. In the case of the intravenous liposomal formulation (AmBisome), the differences between the infected and non-infected tissues are of a higher magnitude than in the case of orally administered AmB (iCo-009).

Visceral leishmaniasis is a deadly parasitic disease caused by obligate intramacrophage protozoans of the Leishmania genus. The World Health Organization estimates the annual death toll to be 50,000, with 500,000 new cases each year. Without treatment, visceral leishmaniasis is inevitably fatal. For the last 70 years, the first line of defense has been pentavalent antimonials; however, increased resistance has brought amphotericin B to the forefront of treatment options. Unfortunately, the difficult route of drug administration, toxicity issues, and cost prevent amphotericin B from reaching the infected population, and mortality continues to rise. Our reformulation of amphotericin B for oral administration has resulted in a highly efficacious antileishmanial treatment that significantly reduces or eradicates liver parasitemia in a murine model of visceral leishmaniasis. This formulation has overcome amphotericin B's significant physicochemical barriers to absorption and holds promise for the development of a self-administered oral therapy for the treatment of visceral leishmaniasis.

The aim of this work was to assess the effect of chronic administration of protonated nanostructured aluminosilicate (NSAS) on the plasma cholesterol levels and development of atherosclerotic lesions in Apolipoprotein (ApoE) deficient mice fed a high cholesterol and high fat diet. Apolipoprotein E (ApoE) deficient mice were divided into the following treatment groups: protonated NSAS 1.4% (w/w), untreated control and 2% (w/w) stigmastanol mixed with high-cholesterol/high-fat diet. Animals were treated for 12 weeks, blood samples were withdrawn every 4 weeks for determination of plasma cholesterol and triglyceride levels. At the end of the study the aortic roots were harvested for assessment of atherosclerotic lesions. NSAS at 1.4% (w/w) and stigmastanol at 2% (w/w) treatment groups showed significant decreases in plasma cholesterol concentrations at all time points relative to the control animals. The lesion sum area in 1.4% (w/w) NSAS and 2% (w/w) stigmastanol groups were significantly less from the control animals. In conclusion, in this study, the effectiveness of chronic administration of protonated NSAS material in the reduction of plasma cholesterol levels and decrease in development of atherosclerotic lesions was demonstrated in Apo-E deficient mice model.

The aim of this work was to assess the ability of aqueous suspensions of surface-modified nanostructured aluminosilicate (NSAS) compounds to reduce the intestinal absorption of cholesterol in a rat model. The rats were divided into 10 treatment groups which included several NSAS compounds at various doses, ezetimibe at 10 mg/kg, stigmastanol at 50 mg/kg, and normal saline. All compounds and controls were independently administered by oral gavage and then a mixture of [(3)H]cholesterol and cold cholesterol in 10% Intralipid(R) was immediately administered orally to the animals. Systemic blood was sampled and the concentration of cholesterol in plasma was determined by means of radioactivity. Protonation of NSAS using an ion-exchange column resulted in significant inhibition of cholesterol absorption relative to the control group (31.5% and 38.6% reduction in absorption of cholesterol for 50 and 100 mg/kg doses, respectively). Other surface-ion modifications of NSAS compounds did not show significant effect on intestinal cholesterol absorption. The inhibition of cholesterol absorption by ezetimibe was superior and by stigmastanol was equal to the effect of protonated NSAS in the doses investigated in this study. In conclusion, protonated NSAS material seems to inhibit significantly the intestinal absorption of dietary cholesterol in a rat model.

Although abdominal surgery is a routine procedure in clinical practice and in preclinical investigation, little is known regarding its effect on the intestinal absorption of drugs. The aim of this study was to investigate the effect of abdominal surgery on the intestinal absorption of highly lipophilic compounds with different absorption mechanisms following oral administration. The 2 compounds that were tested were biopharmaceutical classification system (BCS) class 2 model lipophilic cannabinoid derivatives, dexanabinol and PRS-211,220. Although dexanabinol is mostly absorbed via passive diffusion to the portal blood, PRS-211,220 is absorbed mostly via lymphatic transport. In this work, we compared the absorption of these compounds after abdominal surgery in rat with the absorption data obtained from naïve animals. The outcomes of this investigation showed that the abdominal surgery mostly affected the absorption process on the preenterocyte level, as indicated by the 2-fold increase in the extent of intestinal absorption of dexanabinol, which is a compound with a low degree of intestinal lymphatic transport. However, the lymphatic transport was not affected by the surgical procedure as evident by the absence of change in the extent of absorption of PRS-211,220, which is transported to the systemic circulation mainly by intestinal lymphatics. In conclusion, abdominal surgery can significantly affect the intestinal absorption of lipophilic drugs; however, intestinal lymphatic transport seems to be less affected by the abdominal surgery.

Objective: To develop an oral formulation of Amphotericin B (AmpB) with: (A) medium chain triglycerides, fatty acids and nonionic surfactants as a self-emulsifying drug delivery system (SEDDS); or (B) glyceryl mono-oleate (Peceol) with poly(ethylene glycol) (PEG)-phospholipids., Methods: SEDDS formulations were prepared by simple mixing at 40 degrees C. Peceol/DSPE-PEG-lipid formulations were prepared by solvent evaporation. Parameters evaluated included: miscibility, solubility and emulsion droplet size after incubation in simulated gastric fluid (SGF) or simulated intestinal fluid (SIF) via dynamic light scattering. The stability of AmpB in Peceol/DSPE-PEG was evaluated in SGF and SIF. Phase stability of AmpB in Peceol+/-DSPE-PEG following thermal cycling was evaluated by atomic force microscopy (AFM). Aspergillus fumigatus (2.9-3.45 x 10(7) colony forming units per mL [CFU]) or Candida albicans (3-3.65 x 10(6) CFU per mL) were injected via the jugular vein; 48 h later male albino Sprague-Dawley rats (350-400 g) were administered either a single oral gavage of a Peceol-DSPE/PEG2000-based AmpB (10 mg AmpB/kg and 5 mg AmpB/kg for the Candida albicans study only) twice daily for 2 consecutive days, a single intravenous (i.v.) dose of Abelcet (5mg AmpB/kg), or physiologic saline (non-treated controls; n=9) once daily for 2 consecutive days. Antifungal activity was assessed by organ CFU concentrations and plasma galactomannan levels in the case of A. fumigatus and organ CFU concentrations in the case of Candida albicans. Plasma samples were taken from each animal prior to infection, 48 h after initiation of infection but prior to drug treatment and at the end of the study for plasma creatinine determinations as a measure of renal toxicity., Results: Mean diameter of SEDDS after 30 min in 150 mM NaCl at 37 degrees C was 200-400 nm. However, the Peceol/DSPE-PEG, where PEG MW was 350, 550, 750 or 2000, showed a greater solubilization of AmpB (5 mg/mL) compared to SEDDS formulations (100-500 microg/mL). Upon dispersion in SIF, Peceol/DSPE-PEG formulations generated submicron emulsion particle sizes varying slightly with PEG MW. Stability of the AmpB in Peceol/DSPE-PEG formulations in SGF or SIF was >80% after 2 h, and best for formulations containing DSPE-PEG 750 or 2000 compared to 350, 550 or Peceol only. Monoglyceride-Peceol-DSPE/PEG2000-based oral AmpB treatment significantly decreased total fungal CFU concentrations recovered in all the organs added together by >80% compared to non-treated controls without significant changes in plasma creatinine levels in the A. fumigatus infected rats. In addition, this formulation significantly decreased kidney fungal CFU concentrations by >75% at the 5 mg/kg dose and by >95% at the 10 mg/kg dose compared to non-treated controls without significant changes in the plasma creatinine levels in the Candida albicans-infected rats., Conclusions: Novel lipid-based AmpB oral formulations were prepared that provide excellent drug solubilization, drug stability in simulated gastric and intestinal fluids and antifungal activity without renal toxicity in rats infected with A. fumigatus and C. albicans.

Objectives: The uptake of drugs by chylomicrons is a key element in both intestinal lymphatic transport and postprandial alterations in the disposition profile of lipophilic drugs. The aim of this article was to elucidate the factors that affect this phenomenon., Methods: The degree of association of 22 model lipophilic molecules with rat chylomicrons was assessed and correlated in silico with calculated physicochemical properties. The in-silico model was then validated using an external set of molecules. The uptake by chylomicrons was also compared to the association with a marketed artificial emulsion., Key Findings: The most important physicochemical property that affects the affinity to chylomicrons was found to be LogD7.4; however, a multiparameter model was required to describe properly the uptake process. The in-silico model (R2Y=0.91, R2X=0.91 and Q2=0.82) that was created using a combination of eight molecular descriptors enabled successful prediction of the affinity of the external set of molecules to chylomicrons. The association with the artificial emulsion was statistically different from the uptake by chylomicrons for four (out of nine) molecules., Conclusions: The association of drugs with chylomicrons is a complex process, which involves the lipophilic core as well as surface apoproteins. The in-silico model based on multiple physicochemical properties of the drugs is able to predict successfully the degree of association with chylomicrons.

The introduction of combinatorial chemistry and the development of high-throughput techniques has significantly increased the amount of new active lipophilic molecular entities. As a result, the need for successful delivery of these lipophilic active compounds has also increased. One such method of delivery is through the use of advanced lipid-based drug delivery systems. Traditionally, the majority of reviews of lipid-based drug delivery systems were based on the oral administration field only. This review article focuses on the use of lipids in parenteral, dermal/ transdermal, and ocular drug delivery systems. Specific needs, considerations, approaches, and limitations in lipid-based drug delivery are discussed herein on the basis of the anatomical and physiological diversity of the target organs and systems.

The purpose of this study was to assess the contribution of lymphatics to the systemic bioavailability of macromolecules following SC administration in a rat model. The rat model included continuous lymph collection from the thoracic lymph duct and concurrent serial blood sampling from freely moving animals. A thoracic lymph duct-jugular vein shunt produced by an implanted connective cannula, and maintained during the recovery period, enabled superior rat survival and prevented lymphatic cannula occlusion. The SC absorption of three macromolecules (bovine insulin, bovine serum albumin, and recombinant human erythropoietin alpha) was assessed in comparison to the non-lymph cannulated control group. For all tested molecules, only minimal amounts (less than 3%) of the SC administered dose were detected in the collected lymph. In the rat model, following SC administration, the macromolecules were absorbed mainly through the blood capillaries with minimal contribution of the lymphatic system to systemic bioavailability. The relatively small elevation in the lymphatic concentration, which occurred in all molecules, may be attributed to the redistribution of the molecules from the blood to the interstitial fluid compartment. These findings are important since rodents are commonly used in preclinical evaluation of macromolecular drugs.

Following a high-fat meal, triglyceride-rich lipoproteins (TRL) are assembled in the gut and absorbed via the lymph into the blood circulation, producing a temporal hyperlipidemia. The purpose of this study is to verify the hypothesis that this transient acute postprandial hyperlipidemia affects the pharmacokinetics of lipophilic drugs on both absorption and disposition levels by the same underlying mechanism, namely the association of active lipophilic compounds with TRL in the plasma (disposition) or within the enterocyte (lymphatic transport). This concept was assessed in rats using two model compounds, DDT with high affinity to chylomicrons and diazepam which does not bind to chylomicrons. Oral administration of peanut oil significantly increased the AUC of plasma DDT concentrations following its IV bolus administration in comparison to a water treated group. On the other hand, the AUC of diazepam following IV bolus administration was the same in oil and water treated rats. While DDT is known to have significant lymphatic bioavailability, diazepam has negligible intestinal lymphatic transport (0.014+/-0.004% of a given dose). In conclusion, lipophilic molecules that bind extensively to TRL will be prone to both intestinal lymphatic transport and to post-absorptive changes in disposition (decrease in clearance and volume of distribution) following a high-fat meal.

The aim of this article was to investigate the role of intestinal lymphatic transport in the oral bioavailability of two structurally similar synthetic lipophilic cannabinoids: dexanabinol and PRS-211,220. For this purpose, the long chain triglyceride (LCT) solubility and affinity to chylomicrons ex vivo of both cannabinoids were evaluated. Their oral bioavailability was assessed in rats following administration in a lipid-free and a LCT-based formulation. The intestinal lymphatic transport of these two molecules was also directly measured in a freely moving rat model. LCT solubility of dexanabinol and PRS-211,220 was 7.9+/-0.2 and 95.8+/-5.3mg/g, respectively. The uptake by chylomicrons was moderate (31.6+/-5.2%) and high (66.1+/-2.4%), respectively. The bioavailability of dexanabinol (37%) was not affected by LCT solution, whereas administration of PRS-211,220 in LCT improved the absolute oral bioavailability three-fold (from 13 to 35%) in comparison to the lipid-free formulation. The intestinal lymphatic transport of dexanabinol and PRS-211,220 was 7.5+/-0.8 and 60.7+/-6.8% of the absorbed dose, respectively. In conclusion, despite structural similarity and similar lipophilicity, dexanabinol and PRS-211,220 exhibited a very diverse pattern of oral absorption, and the lymphatic system played quite a different role in the oral bioavailability of these molecules. The low lymphatic transport of dexanabinol is likely driven by relatively lower affinity to chylomicrons and lower LCT solubility.

Cerebrotendinous xanthomatosis (CTX) is a rare neurodegenerative disorder with cholestanol accumulation resulting from mutations in the sterol 27-hydroxylase gene (CYP27A). Conventional treatment includes chenodeoxycholic acid and HMG-CoA reductase inhibitors. Mice with disrupted Cyp27A (Cyp27 KO) do not show elevated cholestanol levels nor develop CTX manifestations. This phenomenon was proposed to be due to murine CYP3A overexpression leading to an alternative pathway for degradation of bile alcohols including cholestanol. Our objective was to examine the influence of CYP3A4 induction on cholestanol elimination in CTX patients. Rifampicin (600 mg/day x 7 days), known to induce the PXR, and thereby to increase CYP3A activity, was used. The degree of CYP3A4 induction was assessed by comparing midazolam pharmacokinetics before and after rifampicin treatment. Cholestanol levels and cholestanol/cholesterol ratios were assayed during the experimental period and compared to a 3 weeks period without treatment. The results show that despite 60% increase in CYP3A4 activity following rifampicin treatment, there is no significant change in cholestanol levels. We conclude that up-regulated expression of CYP3A affects cholestanol elimination in mice differently as compared to its effect in CTX patients. Therefore, CYP3A4 inducers cannot replace chenodeoxycholic acid for the treatment of CTX.

A high-fat meal induces transient hyperlipidemia characterized by elevated triglyceride-rich lipoproteins (TRL) which are composed mainly of chylomicrons. The purpose of this work was to investigate the effect of this transient hyperlipidemia on the pharmacodynamics of lipophilic drugs, using DDT as a model compound since it binds extensively to TRL and has a distinct neurotoxic effect. The postprandial hyperlipidemia in rats was induced by oral administration of peanut oil and was monitored by measurement of plasma triglyceride levels. The control group received water instead of oil. The rats received a continuous intravenous infusion of DDT (10 mg/h) until onset of a predefined pharmacodynamic endpoint (facial muscle tremor). Plasma and brain samples were then obtained and assayed for DDT. Rats with postprandial hyperlipidemia required higher dose of DDT to induce onset of facial muscle tremor. At the pharmacodynamic endpoint, oil treated rats had significantly higher concentrations of DDT in plasma and in the chylomicron fraction, but DDT brain concentrations were the same in both groups. In conclusion, a high-fat meal induces postprandial hyperlipidemia that may significantly alter the pharmacological profile of lipophilic compounds that bind to TRL. This is due to alteration of the distribution characteristics of the lipophilic compound through its association with postprandial lipoproteins. However, this pharmacokinetic phenomenon did not affect the concentration-effect relationship at the site of action in the brain.

Association of a drug with chylomicrons in the enterocyte is an essential step in the lymphatic absorption pathway. In this article, the uptake of lipophilic compounds by chylomicrons ex vivo was compared to the corresponding intestinal lymphatic bioavailability reported in rats in order to elucidate the degree of correlation and to evaluate the utilization of this correlation as a predictive measurement of the lymphatic bioavailability potential of lipophilic drugs. Nine lipophilic compounds (Vitamin D(3), Vitamin E, halofantrine, probucol, diazepam, testosterone, cyclosporin A, benzo[a]pyrene and p,p'-DDT) at a concentration of 1.75 x 10(-6)M were incubated for 1h with chylomicron emulsion separated from rat blood. A strong linear correlation was found between the degree of association of compounds with chylomicrons ex vivo and the lymphatic transport reported in rats (r(2)=0.94, P<0.0001), whereas logP and solubility in long chain triglycerides showed only moderate correlation with lymphatic bioavailability. The linear correlation between the degree of uptake of compounds by isolated chylomicrons and intestinal lymphatic transport suggests that the two processes are governed by similar factors. Thus, the degree of association of lipophilic compounds with isolated chylomicrons can be used as a simple screening model for estimation of intestinal lymphatic transport potential of drug molecules. This approach is important in view of the practical difficulties in direct determination of the lymphatic bioavailability in vivo.

Object-relational mapping (mapping object-oriented software systems to relational databases) remains a challenging undertaking because of the inherent differences between these two technologies. Effective application of this technique in biomedical informatics applications can provide robust, adaptive solutions but requires careful attention to specific issues in the bioscience domain. Mapping from a single, fixed, stably defined object to a table is straightforward, and several standard patterns have been described for mapping inheritance trees to tables. However, mapping becomes more problematic when it involves complex, evolving inheritance trees, sparse, dynamic attributes, or associations and aggregations whose requirements and definitions change frequently during development. All of these seem to be inherent, inescapable facts in the development of biological information systems. Here we describe modifications of the basic patterns of object-relational mapping that are being applied to two systems in ongoing collaborations between biomedical investigators and informaticians. In this approach a small set of objects for each entity collaborates to provide the usual required services of an object-relational implementation including query capacity, transaction management, object relationships (associations, aggregation, composition, many-many relations) and persistence functions. This set of objects also allows dynamic attributes for a selected set of entities by merging an entity-attribute-value approach for data storage with the object-relational approach to managing relationships among objects. Templates are used for class generation to allow flexibility and synchronization of schema/object during design evolution, facilitating changing persistence strategies across the whole system for testing and development. The entity-attribute-value details as well as all database access are encapsulated in a persistence layer which would allow change of this layer to a conventional storage model, if the design evolution is stabilized or for performance requirements, with no change to the domain or presentation layers.

Access to timely decision support information is critical for delivery of high-quality medical care. Transformation of clinical knowledge that is originally expressed in the form of a guideline to a computable format is one of the main obstacles to the integration of knowledge sharing functionality into computerized clinical systems. The Guideline Element Model (GEM) provides a methodology for such a transformation. Although the model has been used to store heterogeneous guideline knowledge, it is important to demonstrate that GEM markup facilitates guideline implementation. This report demonstrates the feasibility of implementation of GEM-encoded guideline recommendations using Apache Group s Cocoon Web Publishing Framework. We further demonstrate how XML-based programming allows for maintaining the separation of guideline content from processing logic and from presentation format. Finally, we analyze whether the guideline authors original intent has been sufficiently captured and conveyed to the end user.

Implementation of practice guidelines refers to the creation of strategies and systems to operationalize the knowledge and recommendations set forth by guideline developers. We describe an approach to guideline implementation that makes direct use of the guideline document as a knowledge base. The Guideline Elements Model (GEM) provides an XML-based guideline document model that facilitates implementation of guidelines. Knowledge extraction using GEM requires document markup rather than programming and can promote authenticity and consistent knowledge encoding. Knowledge customization for the local enterprise requires addition of meta-information to pertinent components of the GEM hierarchy in a design database. GEM provides an audit trail to track local adaptation. Knowledge integration with patient data can be promoted using information management services. A design goal is to devise a system that can be applied by local clinical domain experts, quality assurance experts, and information systems programmers without requiring trained informaticians and knowledge engineers to serve as intermediaries